The overall objective of this project is to examine in a direct way the interactions between the heme and the globin in hemoglobin. This will be accomplished by preparing a series of chemically modified hemes, in which either the peripheral substituents or the nature of the sixth ligand are altered in such derivatives, the alterations in structure will be determined by x-ray crystallographic techniques, and correlated with the alterations in functional properties observed by such techniques as ligand equilibria and (in collaboration with Prof. Quentin Gibson) rapid kinetics. A complete structure determination of hemoglobin reconstituted with 2,4-dibromoheme by x-ray difference Fourier techniques is clearly feasible, and a similar study of native hemoglobin reacted with thiocyanate may also be possible. It is extremely desirable to compare the former with the structure of the corresponding methyl derivative, as much of our interpretation of kinetic data rests on the close isomorphism of this pair. Attempts will be made to recrystallize the methyl derivative, by harvesting the showers of microcrystals already obtained, dissolving them in dilute buffer, and modifying the crystallization conditions to diminish the number of nuclei. With the development of the Cornell synchrotron radiation facility, a major effort will go into designing and constructing the apparatus necessary to conduct EXAFS and single crystal diffraction experiments, in collaboration with the synchrotron staff. Many of the hemoglobins we have been studying are ideal for investigation by EXAFS techniques; the location of the iron atoms in single crystals can be determined exceptionally accurately by anomalous scattering measurements.